Project 1

项目1

• 批准号: 8744859
• 负责人: Robert Ros
• 金额: $ 15.01万
• 依托单位: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8744859
• 关键词: Atomic Force Microscopy; Biological Assay; Cancer Biology; Cells; Constitution; Data; Development; Elasticity; Extracellular Matrix; FarGo; Finite Element Analysis; Fluorescence; Gel; Image; Lasers; Laws; Life; Luciferases; Malignant - descriptor; Malignant Neoplasms; Maps; Measures; Mechanical Stress; Mechanics; Methods; Microscopy; Molecular; Morphology; Optics; Pathway interactions; Phase-Contrast Microscopy; Population; Property; Reporter; Resolution; Sampling; Scanning; Shapes; Signal Pathway; Signal Transduction; Solutions; Spectrum Analysis; Stimulus; Surface; Techniques; Technology; Time; aqueous; base; cancer cell; nanometer; nanoscale; novel; novel strategies; physical property; physical science; response; tumorigenic; vector

): Cancer development is associated with changes in the mechanical properties of cells as well as the extracellular matrix. Few facts are known about the interplay of cells and the extracellular matrix (ECM) on the molecular level in respect to the transformation from a normal to a malignant cell. The objective of this proposed project is to develop a radically new approach to investigate quantitatively the mechanical properties of cells and ECM and to correlate those with signaling pathways. Cells will be embedded in true 3D-matrixes and a method based on the combination of atomic force microscopy (AFM) and the optical microscopy techniques, laser scanning confocal, wide-field fluorescence, and phase contrast microscopy will be applied to extract mechanical volume properties ofthe cells and the surrounding matrix with nanometer resolution. Because ofthe heterogeneous constitution, on the nanometer scale, ofthe cells as well as the matrix, new analysis methods based on finite element analysis have to be developed and applied in order to extract quantitative data to create 3D-elasticity maps ofthe samples. Using this novel method, the influence of local induced mechanical stress on the shape, surface morphology, and the signaling pathways ofthe cells will be investigated. In general, this ansatz enables us to correlate the physical properties, malignant transformations, and molecular tumorigenic signaling pathways. The proposed technique is a single cell method, which allows for the exploration of inhomogeneities in a cell population. The specific aims of this proposed project are: 1) Development of mechanical nanotomography of cells embedded in true 3D-matrixes for quantitative analysis of local mechanical properties. 2) Development of gel encapsulation of living cells compatible with mechanical nanotomography. The cell response to encapsulation in 3D-matrixes of different stiffness and composition will be measured. This method will be applied to cells embedded in matrixes mimicking ECM. 3) Development of a method based on combined AFM/optical microscopy to probe the response ofthe embedded cells to external mechanical stimuli.

）：癌症发展与细胞的机械性能以及细胞外基质的变化有关。关于从正常细胞到恶性细胞的转化，很少有关于细胞的相互作用和细胞外基质（ECM）的事实。该提出的项目的目的是开发一种彻底的新方法，以定量研究细胞和ECM的机械性能，并将其与信号通路相关联。细胞将嵌入真实的3D-Matrix和基于原子力显微镜（AFM）和光学显微镜技术的组合，激光扫描共聚焦，宽视野荧光和相对比显微镜的组合将应用于提取机械体积的特性带有纳米分辨率的细胞和周围矩阵。由于在纳米尺度上，细胞和基质的异质构成，必须开发并应用基于有限元分析的新分析方法，以提取定量数据以创建样品的3D弹性图。使用这种新方法，将研究局部诱导的机械应力对细胞的形状，表面形态和信号通路的影响。通常，这种安萨斯使我们能够将物理特性，恶性转化和分子肿瘤信号通路相关联。提出的技术是一种单细胞方法，它允许探索细胞种群中的不均匀性。该提出的项目的具体目的是：1）开发嵌入真实3D-矩阵中的细胞的机械纳米摄影，以定量局部机械性能分析。 2）与机械纳米摄影兼容的活细胞的凝胶封装的开发。将测量对不同刚度和组成的3D-矩阵封装的细胞反应。该方法将应用于模仿ECM的基质中的细胞。 3）开发基于AFM/光学显微镜组合的方法，以探测嵌入式细胞对外部机械刺激的响应。